Allogeneic hematopoietic cell transplant (HCT) has improved survival of patients with leukemia, but relapse remains the major cause of treatment failure. The infusion of donor lymphocytes to treat relapse after HCT has formally demonstrated benefits of lymphocyte-mediated anti-leukemic activity, but the polyclonal T cells recognize proteins expressed by many host tissues as well as leukemia, resulting in morbidity/mortality from graft-versus-host disease. Thus, generating and administering T cells specific for proteins preferentially expressed by leukemic cells has potential to not only impact survival but provide a platform for developing T cell therapy as a more effective, less toxic therapeutic modality. WT1, a pro-oncogenic transcription factor with very limited expression in nonrial adult tissues, is detected at high levels in leukemic cells and many other malignancies. We have shown that WTI-specific CD8 T cells, generated from HCT donors, expanded ex vivo, and transferred into relapsed patients, can exhibit anti-leukemic activity with no evidence of toxicity to normal tissues, but most patients progress in part due to failure to achieve large magnitude T cell responses of high avidity that can persist to eradicate leukemia. Similar obstacles have been observed with WT1 vaccines. These problems may be addressable by employing lentiviral vectors to insert high affinity TCR genes isolated from a WT1-specific CD8 T cell done into large numbers of donor memory CD8 T cells with limited ex vivo expansion. The proposed studies will: 1) evaluate in a Phase l/ll trial safety and potential efficacy for preventing relapse in HCT patients at high risk for recurrence of infusing donor-derived EBV specific CD8 T cells transduced to express a high affinity TCR specific for WT1; 2) evaluate in a Phase l/ll trial potential limitations to anti-leukemic activity of infusions of these transduced WTI-specific CDS T cells into patients who have relapsed after HCT; and 3) generate and screen for safety/activity a panel of mutated human TCRs with a range of increased affinities for WT1 as a means to overcome obstacles to efficacy in future trials for leukemia and other tumors. These studies using novel reagents/methods should impact fields of T cell and gene therapy and demonstrate the potential of gene-modified T cells as therapeutic reagents.